Method of soft tissue augmentation

ABSTRACT

Particles according to the invention are made of a viscoelastic medium, are injectable gel particles, and have a size, when subjected to a physiological salt solution, in the range of from 1 to 5 mm. The particles are useful in a soft tissue augmentation implant. The implant comprises particles of a viscoelastic medium, wherein a major volume of the particles are injectable gel particles according to the invention. The implant is useful in a method of soft tissue augmentation in a mammal, including man, comprising subepidermal administration at a site in said mammal where soft tissue augmentation is desirable, of an implant according to the invention.

This is a Division of application Ser. No. 11/090,141 filed on Mar. 28,2005, which claims the benefit under 35 U.S.C. §119(e) of U.S.Provisional Patent Application No. 60/560,258, filed Apr. 8, 2004. Thedisclosure of the prior applications is hereby incorporated by referenceherein in their entirety.

FIELD OF THE INVENTION

The present invention relates to the fields of esthetics and plasticsurgery, including cosmetic and reconstructive surgery. Morespecifically, the invention is concerned with a method of soft tissueaugmentation in a mammal, including man. Moreover, the invention isdirected to use of particles of a viscoelastic medium for themanufacture of a medicament for therapeutic soft tissue augmentation ina mammal, including man. The invention is also concerned with particlesof a viscoelastic medium, production thereof, and use thereof in animplant.

BACKGROUND TO THE INVENTION

An implant material that is useful for soft-tissue augmentation shouldideally be capable of providing adequate and sustained aesthetic and/ortherapeutic correction without migration and displacement;natural-looking and non-palpable; easy to administer and, if necessary,remove; non-immunogenic; and devoid of chronic inflammatory reactions(Krauss M C, Semin Cutan Med Surg 1999; 18: 119-128). As a soft-tissueaugmentation material, hyaluronic acid (a naturally occurringpolysaccharide) has low immunogenic potential, being chemicallyhomogenous across all species and tissues (Larsen N E et al., J BiomedMater Res 1993; 27: 1129-1134). Stabilization (or cross-linking) of thehyaluronic acid molecule improves its resistance to enzymaticdegradation without compromising its biocompatibility, while the use ofa non-animal source reduces the likelihood of antigenic contaminationand subsequent hypersensitivity reactions (Friedman et al., DermatolSurg 2002; 28: 491-4).

Non-animal stabilized hyaluronic acid (NASHA), U.S. Pat. No. 5,827,937,may be produced from a highly purified hyaluronic acid preparationobtained by bacterial fermentation. Various NASHA preparations ofdifferent particle size (Restylane® Perlane, Restylane®, Restylane® FineLines and Restylane® Touch, all from Q-Med AB, Uppsala, Sweden) havebeen developed as dermal fillers for facial soft-tissue augmentation.Clinical studies indicate that known NASHA gels are effective inaugmenting lips (Bousquet M-T and Ågerup B, Oper Techniques OcuplastOrbit Reconstruct Surg 1999; 2: 172-176) and correcting facial wrinklesand folds (Olenius M. Aesth Plast Surg 1998; 22: 97-101; Duranti F etal., Dermatol Surg 1998; 24:1317-25; Narins RS et al., Dermatol Surg2003; 29: 588-95; Carruthers J and Carruthers A, Dermatol Surg 2003; 29:802-9), and that they offer a more durable aesthetic improvement thanbovine collagen or hylan B. The extensive clinical experience gainedfrom their intradermal use in some 1.5 million facial cosmeticprocedures confirms their safety.

It is recommended that RESTYLANE Touch (˜500,000 particles/ml, meanparticle size 0.2 mm) should be injected in the upper part of thedermis; RESTYLANE (˜100,000 particles/ml, mean particle sizeapproximately 0.4 mm) should be injected in the mid-part of the dermis;and RESTYLANE Perlane (˜10,000 particles/ml, mean particle sizeapproximately 0.8 mm) should be injected in the deep layer of the dermisand/or the surface layer of the subcutis.

Some known soft-tissue augmentation treatments involving implantation ofviscoelastic materials occasionally suffer from the drawback that theimplant, or part thereof, migrates away from the desired site oftreatment. Another problem with some known tissue augmentationtreatments involving implantation of viscoelastic materials is that theimplant is displaced from the desired site of treatment. Implantmigration and displacement are disadvantageous for the patient, sincethey may impair the cosmetic and/or therapeutic outcome of the treatmentand may impede removal of the implant, if this is desired.

SUMMARY OF THE INVENTION

It is an object of the present invention to provide an implantationmaterial suitable for soft tissue augmentation which overcomes drawbackswith known implantation materials. It is also an object of the presentinvention to provide a method of producing such an implantationmaterial.

It is another object to provide an implant suitable for soft tissueaugmentation comprising such an implantation material which overcomesdrawbacks with known implants. It is a further object of the presentinvention to provide an implant suitable for soft tissue augmentationcomprising such an implantation material which is readily removable ifdesired.

It is another object of the present invention to provide a method ofsoft tissue augmentation in a mammal, including man, which overcomesdrawbacks with known methods. It is one object of the present inventionto provide a method of soft tissue augmentation in a mammal, includingman, comprising subepidermal administration of an implant, which avoidsor diminishes undesired migration of the implant from the desired siteof implantation. It is one object of the present invention to provide amethod of soft tissue augmentation in a mammal, including man,comprising subepidermal administration of an implant, which avoids ordiminishes displacement of the implant from the desired site ofimplantation.

It is also an object of the present invention to provide use of aviscoelastic medium for the manufacture of a medicament for therapeuticsoft tissue augmentation in a mammal, including man.

For these and other objects that will be evident from the followingdisclosure, the present invention provides particles of a viscoelasticmedium, which are injectable gel particles having a size, when subjectedto a physiological salt solution, in the range of from 1 to 5 mm.

The invention is based on the finding that subepidermal administrationof an implant comprising gel particles made of a viscoelastic mediumwhich are considerably larger than previously used in implants made ofviscoelastic media are useful in avoiding migration and/or displacementof the implant, or part thereof, from the desired site of soft tissueaugmentation. Moreover, the limited displacement of the implant incombination with the considerable particle size facilitates easy removalof the implant, if desired.

In certain preferred particles according to the invention, said size isin the range of from 1 to 2.5 mm. In other preferred particles accordingto the invention, said size is in the range of from 2.5 to 5 mm.

In preferred particles according the invention, said viscoelastic mediumis selected from the group consisting of polysaccharides and derivativesthereof. In more preferred particles according the invention, saidviscoelastic medium is selected from stabilized glycosaminoglycans andderivatives thereof. In certain preferred particles according theinvention, said viscoelastic medium is selected from the groupconsisting of stabilized hyaluronic acid, stabilized chondroitinsulfate, stabilized heparin, and derivatives thereof.

In preferred particles according the invention, said viscoelastic mediumis selected from the group consisting of cross-linked hyaluronic acidand derivatives thereof. In particularly preferred particles accordingthe invention, the concentration of said viscoelastic medium in said gelparticles, when subjected to a physiological salt solution, is in therange of from 5 to 100 mg/ml.

Preferred particles according to the invention are injectable through a20 gauge or larger needle by application of a pressure of 15-50 N.

According to another aspect of the invention, there is provided a methodof producing injectable gel particles of a viscoelastic medium,comprising the steps of: (i) manufacturing a gel with a desiredconcentration of said viscoelastic medium; and (ii) mechanicallydisrupting said gel into gel particles having a size, when subjected toa physiological salt solution, in the range of from 1 to 5 mm.

According yet another aspect of the invention, there is provided a softtissue augmentation implant comprising particles of a viscoelasticmedium, wherein a major volume of said particles are injectable gelparticles having a size, when subjected to a physiological saltsolution, in the range of from 1 to 5 mm. In preferred embodiments ofthe implant, said size is in the range of from 1 to 2.5 mm. In otherpreferred embodiments of the implant, said size is in the range of from2.5 to 5 mm.

According to one aspect of the invention, there is provided a method ofsoft tissue augmentation in a mammal, including man, comprisingsubepidermal administration at a site in said mammal where soft tissueaugmentation is desirable, of an implant comprising injectable gelparticles of a viscoelastic medium, a major volume of said particleshaving a size, when subjected to a physiological salt solution, in therange of from 1 to 5 mm. In preferred embodiments of the methodaccording to the invention, said administration is selected from thegroup consisting of subcutaneous administration, submuscularadministration and supraperiostal administration.

In certain methods according to the invention, said size is in the rangeof from 1 to 2.5 mm. In a preferred embodiment of this method, said siteof soft tissue augmentation is selected from facial tissue and othertissues covered by exposed skin. In other methods according to theinvention, said size is in the range of from 2.5 to 5 mm.

In preferred methods according to the invention, said administration isa selected from the group consisting of single administration andmultiple-layer administration.

According to another aspect of the invention, there is providedinjectable gel particles according to the invention for use as amedicament. There is also provided an injectable soft tissueaugmentation implant comprising injectable gel particles according tothe invention for use as a medicament.

According to yet another aspect of the invention, there is provided useof injectable gel particles of a viscoelastic medium according to theinvention, a major volume of said particles having an average size, whensubjected to a physiological salt solution, in the range of from 1 to 5mm, for the manufacture of a medicament for therapeutic soft tissueaugmentation in a mammal, including man, wherein said medicament issuitable for subepidermal administration according to the invention at asite in said mammal where therapeutic soft tissue augmentation isdesirable.

DETAILED DESCRIPTION OF THE INVENTION

According to one aspect of the invention, there is provided particles ofa viscoelastic medium, which are injectable gel particles having a size,when subjected to a physiological salt solution, in the range of from 1to 5 mm. The particles are useful in a soft tissue augmentation implantcomprising particles of a viscoelastic medium, wherein a major volume ofsaid particles are injectable gel particles according to the invention,said gel particles having a size, when subjected to a physiological saltsolution, in the range of from 1 to 5 mm. The implant, in turn, isuseful in a method of soft tissue augmentation in a mammal, includingman, comprising subepidermal administration at a site in said mammalwhere soft tissue augmentation is desirable, of an implant comprisinginjectable gel particles of a viscoelastic medium, a major volume ofsaid particles having a size, when subjected to a physiological saltsolution, in the range of from 1 to 5 mm.

The term “soft tissue augmentation”, as used herein, refers to any typeof volume augmentation of soft tissues, including, but not limited to,facial contouring (e.g. more pronounced cheeks or chin), correction ofconcave deformities (e.g. post-traumatic, HIV associated lipoatrophy)and correction of deep age-related facial folds. Thus, soft tissueaugmentation may be used solely for cosmetic purposes or for medicalpurposes, such as following trauma or degenerative disease.

The term “soft tissue”, as used herein, refers to tissues that connect,support, or surround other structures and organs of the body. Softtissue includes muscles, fibrous tissues and fat.

The method according to the invention may be performed in any mammal,including man. It is preferred that that the method is performed in ahuman subject.

The terms “subepidermal administration” or “subcuticularadministration”, as used herein, refer to administration beneath theepidermis of the skin, including administration into the dermis,subcutis or deeper, such as submuscularly or into the periosteum whereapplicable (in the vicinity of bone tissue.

Administration may be performed in any suitable way, such as viainjection from standard cannulae and needles of appropriate sizes. Theadministration is performed where the soft tissue augmentation isdesired, such as the chin, cheeks or elsewhere in the face or body.

The term “implant”, as used herein, refers widely to any type ofimplanted or implantable foreign object or material. Implants alsoinclude objects or materials that are nearly identical to non-foreignobjects or materials. The implant according to the invention is notlimited to any particular shape. The final shape of the implant in thebody is decided by the skilled man from the purpose of the treatment.

By the term “viscoelastic medium”, as used herein, is meant a mediumthat exhibits a combination of viscous and elastic properties.Specifically, the viscoelastic medium according to the invention isinjectable through a 20 gauge or larger needle, such as a 10-20 gaugeneedle, by application of a pressure of 15-50 N. In particular, themedium, or an implant or a medicament comprising the medium, is suitablefor subepidermal injection into a human in need thereof at a desiredsite.

Viscoelastic media according to the invention include gels, dispersions,solutions, suspensions, slurries and mixtures thereof. It is preferredthat the medium is present as a dispersion of gel or gel-like particles.In a preferred embodiment, the implant according to the inventionconsists of 1-5 mm large particles of one or more viscoelastic mediadispersed in a physiological salt buffer or a suitable physiologicalsalt solvent. In another preferred embodiment, the implant furthercomprises other suitable additives, such as local anesthetics,anti-inflammatory drugs, antibiotics and other suitable supportivemedications, e.g. bone growth factors or cells. Optionally, there mayalso be included a viscoelastic medium, which may be the same ordifferent, which is not present as particles or as particles of a sizesmaller than 0.1 mm.

It goes without saying that the size of the gel particles according tothe invention is dependent upon the ionic strength of the buffer,solution or carrier that is included in and/or surrounding the gelparticles. Throughout this specification, given particle sizes assumephysiological conditions, particularly isotonic conditions. It shall benoted that, while it is preferred that the gel particles contain and aredispersed in a physiological salt solution, it is contemplated that thegel particles according to the invention can temporarily be brought todifferent sizes by subjecting the gel particles to a solution of anothertonicity. Particles that are within the scope of this invention exhibita particle size within the given ranges under physiological conditions,e.g. when implanted subepidermally in the body or when subjected to aphysiological, or isotonic, salt solution, i.e. a solution with the sametonicity as the relevant biological fluids, e.g. isoosmotic with serum.

Thus, the viscoelastic medium according to the invention is present atleast as gel particles or gel-like particles. A major volume, or morethan 50% (v/v), of the particles have a size of at least 1 mm,preferably in the range of 1-5 mm in the presence of a physiologicalsalt solution. In preferred embodiments, more than 70% (v/v), preferablymore than 90% (v/v), of the particles are within the given size limitsunder physiological conditions.

As used herein, a physiological, or isotonic, solution is a solutionhaving an osmolarity in the range of 200-400 mOsm/l, preferably 250-350mOsm/l, more preferably approximately 300 mOsm/l. For practicalpurposes, this osmolarity is easily achieved by preparation of a 0.9%(0.154 M) NaCl solution.

A suitable way of obtaining a desired particle size involves producing agel made of a viscoelastic medium at a desired concentration andsubjecting the gel to physical disruption, such as mincing, mashing orallowing the gel to pass through a filter with suitable pore size. Theresulting gel particles are dispersed in a physiological salt solution,resulting in a gel dispersion or slurry with particles of desired size.

Another aspect of the invention is the density, or hardness, of the gelparticles. The gel particle density can readily be regulated byadjustment e.g. of the concentration of the viscoelastic medium and theamount and type of cross-linking agent, if any. Thus, harder particlescan be achieved by a higher concentration of the viscoelastic medium inthe gel, and thereby in the resulting gel particles. Harder particlesare generally less viscoelastic and have a longer half-life in vivo thansofter particles. For use in the present invention, it is critical thatthe particles retain enough viscoelastic properties so that they arestill injectable.

In a preferred embodiment of the invention, the implant is atwo-component composition, consisting of softer gel particles mixed withharder gel particles. The gel particles may be made of the same ordifferent viscoelastic media. The resulting mixture of gel particlescombines desirable properties of softness/hardness for use in softtissue augmentation and long durability in vivo.

Administration of the implant employing the method according to theinvention prevents or diminishes migration and/or displacement of theimplant, which comprises or consists of the 1-5 mm large particles underphysiological conditions. A further advantage of the invention is thatthe large size of the particles in combination with the prevented ordiminished migration facilitate easy removal of the implant comprisingthe particles, should it be desired for some reason.

In a preferred embodiment of the invention, the particles have a size inthe range of from 1 to 2.5 mm, such as from 1.5 to 2 mm, in the presenceof a physiological salt solution. These particles are suitable foradministration to subcutaneous, submuscular or supraperiostal tissue. Inparticular, they are suitable for administration to tissues covered byskin that is exposed in public, such as facial tissue, since theparticles and needles that are suitable for this particle size range arenot likely to cause bruises or other discolorations. In a preferredembodiment, these particles are administered to deep subcutaneous or tosubmuscular/supraperiostal tissue, optionally in more than one layer.Deep subcutaneous or submuscular/supraperiostal administration furtherprevents or diminishes migration of the particles away from the desiredsite. According to this embodiment, a major volume, or more than 50%(v/v), preferably more than 70% (v/v), more preferably more than 90%(v/v), of the particles are within the given size limits underphysiological conditions.

In another embodiment of the invention, the particles have a size in therange of from 2.5 to 5 mm, such as from 3 to 4 mm, in the presence of aphysiological salt solution. Implants comprising such particles furtherprevents or diminishes migration of the particles away from the desiredsite. According to this embodiment, a major volume, or more than 50%(v/v), preferably more than 70% (v/v), more preferably more than 90%(v/v), of the particles are within the given size limits underphysiological conditions.

Particle size may be determined in any suitable way, such as by laserdiffraction, microscopy, filtration, etc, and is decided by the longestdistance between two ends of the particle. The specific shape of the gelparticles is not critical. For spherical particles, the diameter equalsthe size for this purpose. The size range may be regulated by mechanicaldisruption, such as mincing, mashing, filtration, etc, of a gel of asuitable concentration of the desired viscoelastic medium.

Viscoelastic media according to the invention include, without beinglimited thereto, polysaccharides and derivatives thereof. Suitableviscoelastic media include stabilized starch and derivatives thereof.Suitable viscoelastic media can also be selected from stabilizedglycosaminoglycans and derivatives thereof, such as stabilizedhyaluronic acid, stabilized chondroitin sulfate, stabilized heparin, andderivatives thereof. Suitable viscoelastic media also include stabilizeddextran and derivatives thereof, such as dextranomer. The viscoelasticmedium may also be a combination of two or more suitable viscoelasticmedia.

By the term “stabilized”, as used herein, is meant any form of chemicalstabilization that, under physiological conditions, renders thestabilized compound more stable to biodegradation that the parentcompound. Without being limited thereto, stabilized compounds includecross-linked compounds and partially cross-linked compounds.

By the term “derivative” of a polysaccharide, as used herein, is meantany suitable derivative thereof, including cross-linked polysaccharidesand substituted polysaccharides, such as sulfated polysaccharides.

Viscoelastic media according to the invention are biocompatible, sterileand present as particles that are readily injectable through standardneedles used in medicine, such as 20 gauge or larger needles, preferably10-20 gauge needles. It is preferable that the viscoelastic medium is ofnon-animal origin. Advantageously, the media according to the inventionare stable, but not permanent, under physiological conditions. Accordingto an embodiment of the invention, at least 70%, preferably at least90%, of the viscoelastic medium remains for at least two weeks in vivo,more preferably between two weeks and two years. The viscoelastic mediumaccording to the invention is preferably degraded after five years ormore in vivo. The term “degraded” implies that less than 20%, preferablyless than 10%, of the medium remains in the body.

The viscoelastic medium according to the invention is more resistant tobiodegradation in vivo than natural hyaluronic acid. The prolongedpresence of the stable viscoelastic substance is advantageous for thepatient, since the time between treatments is increased.

A preferable viscoelastic medium according to the invention iscross-linked hyaluronic acid and derivatives thereof. One type ofsuitable cross-linked hyaluronic acid is obtainable by cross-linking ofhyaluronic acid, optionally non-animal, using the method of U.S. Pat.No. 5,827,937.

In brief, said method involves forming an aqueous solution of a watersoluble, cross-linkable polysaccharide; initiating a cross-linking ofthe polysaccharide in the presence of a polyfunctional cross-linkingagent; sterically hindering the cross-linking reaction from terminatingbefore gelation occurs, whereby an activated polysaccharide is obtained;and reintroducing sterically unhindered conditions for the activatedpolysaccharide so as to continue the cross-linking thereof up to aviscoelastic gel.

The cross-linking agent to be used in connection with this particularmethod is any previously known cross-linking agent useful in connectionwith polysaccharides, consideration being taken to ensure that thebiocompatibility prerequisites are fulfilled. Preferably, however, thecross-linking agent is selected from the group consisting of aldehydes,epoxides, polyaziridyl compounds, glycidyl ethers and divinylsulfones.Of these, glycidyl ethers represent an especially preferred group, ofwhich 1,4-butanediol diglycidyl ether can be referred to as a preferredexample.

In this particular method, the initial cross-linking reaction in thepresence of a polyfunctional cross-linking agent can be performed atvarying pH values, primarily depending on whether ether or esterreactions should be promoted.

An example of a preferred viscoelastic medium is non-animal stabilizedhyaluronic acid, commercially available from Q-Med AB, Uppsala, Sweden.

When the injectable medium is a hyaluronic acid medium, the hyaluronicacid concentration is 5 mg/ml or higher. It is preferred that thehyaluronic acid concentration is in the range of 5-100 mg/ml, morepreferred 10-50 mg/ml, such as approximately 20 mg/ml.

The cross-linked hyaluronic acid is present as particles or beads of anyform. According to this embodiment, a major volume, or more than 50%(v/v), preferably more than 70% (v/v), more preferably more than 90%(v/v), of the particles are at least 1 mm in size, preferably in therange of 1-5 mm. As outlined above, a preferred embodiment involvesparticles in the range of 1-2.5 mm, preferably 1.5-2 mm. Anotherpreferred embodiment involves particles in the range of 2.5-5 mm,preferably 3-4 mm.

A suitable way of obtaining a desired particle size involves producing agel made of cross-linked hyaluronic acid at a desired concentration andsubjecting the gel to physical disruption, such as mincing, mashing orallowing the gel to pass through a filter with suitable particle size.The resulting gel particles are dispersed in a physiological saltsolution, resulting in a gel dispersion or slurry with particles ofdesired size.

Another aspect of the invention is the density, or hardness, of theparticles. Using the manufacturing method of the invention, thecross-linked hyaluronic acid particle density can readily be regulatedby adjustment of the concentration of the viscoelastic medium and theamount and type of cross-linking agent. Thus, harder particles can beachieved by a higher concentration of the viscoelastic medium in thegel, and thereby in the resulting gel particles. Harder particles aregenerally less viscoelastic and have a longer half-life in vivo thansofter particles. Useful hyaluronic acid concentrations yielding gelparticles of varying hardness are e.g. 20, 25, 40, 50 and 100 mg/ml. Foruse in the present invention, it is critical that the particles retainenough viscoelastic properties so that they are still injectable asdiscussed above.

In a preferred embodiment of the invention, softer gel particles, e.g.15-22 mg/ml cross-linked hyaluronic acid, are mixed with harder gelparticles, e.g. 22-30 mg/ml cross-linked hyaluronic acid. The resultingmixture of gel particles combines desirable properties ofsoftness/hardness for use in soft tissue augmentation and longdurability in vivo.

According to the invention, the viscoelastic medium is administered,preferably injected, under the epidermis in any suitable way. By way ofexample, a dermal incision can be made with a scalpel or a sharpinjection needle to facilitate transdermal insertion of a larger cannulafor administration of the implant according to the invention at thedesired site. It is preferred that the administration is performedsubcutaneously, submuscularly or supraperiostally.

The implant, consisting of particles of a viscoelastic medium andoptionally other suitable ingredients, may be administered as a singlealiquot or as layers of multiple aliquots. Optionally, the viscoelasticmedium may be replaced, refilled or replenished by a subsequentinjection of the same or another viscoelastic medium. The injectedvolume is determined by the desired augmentation. In a typical tissueaugmentation, a volume in the range of 1-500 ml is injected, dependingon the purpose and the treated tissue.

According to another aspect of the invention, there is provided a noveluse of particles of a viscoelastic medium according to the invention, amajor volume of said particles having an average size in the range offrom 1 to 5 mm, for the manufacture of a medicament for therapeutic softtissue augmentation in a mammal, including man, wherein said medicamentis suitable for subepidermal administration according to the inventionat a site in said mammal where therapeutic soft tissue augmentation isdesirable.

According to this aspect, it is preferred that the administration isperformed subcutaneously, submuscularly or supraperiostally. Thediscussion hereinabove regarding suitable particle sizes applies alsofor this aspect of the invention.

As used herein, the term “therapeutic” involves any kind of preventive,alleviating or curative treatment. Without being limited thereto, thisaspect of the invention encompasses that the medicament is forreconstructive augmentation resulting from a medical condition and ispart of a medical treatment of the condition. Thus, the therapeutic useis distinguishable from the non-medical, or cosmetic, use in that theyare directed to different patient groups. Specifically, the therapeuticuse is solely directed to patients in need of reconstructiveaugmentation resulting from a medical condition and constitutes a partof a medical treatment of this condition in these patients.

Without being limited thereto, the present invention will in thefollowing be further illustrated by way of examples.

EXAMPLES Example 1

Preparation of Gel Particles of Non-Animal Stabilized Hyaluronic Acid

As previously exemplified in e.g. U.S. Pat. No. 5,827,937, 10 g ofhyaluronic acid prepared by fermentation of Streptococcus was dissolvedin 100 ml of 1% NaOH, pH>9. Cross-linking agent in the form of1,4-butanediol diglycidyl ether was added to a concentration of 0.2%.The solution was incubated at 40° C. for 4 h.

The incubated solution was diluted with an acidic water solution toreach neutral pH under mixing, yielding a final hyaluronic acidconcentration of 20 mg/ml, and again incubated for 12 h at 70° C. Theviscoelastic slurry that resulted from this second incubation was thencooled to room temperature and mashed to its final particle size,approximately 1.5-2 mm.

Example 2

Cheek and Chin Augmentation

Materials

A clear, colorless, viscoelastic gel consisting of non-animal stabilizedhyaluronic acid (20 mg/ml) dispersed in physiological saline solution.The gel is obtainable e.g. by the method of example 1. The sterilizedstudy material (2 ml) was supplied in a 3 ml glass syringe and wasinjected subcutaneously and/or supraperiostally using a sterilized 16G×7 or 9 cm Coleman infiltration cannula with a blunt tip (Byron MedicalInc., Tucson, Ariz., USA).

Patient Selection and Study Design

Adult outpatients (>18 years of age) of either gender seeking cheekand/or chin augmentation therapy for aesthetic purposes. For studyinclusion, patients were required to agree to abstain from othercosmetic procedures (e.g., further augmentation therapy, botulinum toxininjections, laser or chemical skin resurfacing or facelift procedures)for the duration of the study. Patients who had undergone facial tissueaugmentation therapy or laser/chemical peeling procedures within theprevious 6 months or aesthetic facial surgery within the previous 12months were excluded from the study, In addition, patients presentingwith active skin disease or inflammation affecting the intendedtreatment area, those with known allergy/hypersensitivity to localanaesthetics or previous adverse reactions to NASHA, and those currentlytaking anticoagulant or antiplatelet drugs were excluded fromparticipation. The use of anticoagulants, aspirin and non-steroidalanti-inflammatory drugs was prohibited until the injection site hadcompletely healed.

Injection Technique

The treatment area was cleaned with an antiseptic solution and, if localanaesthesia was required, lidocaine (0.5 or 1.0%)/adrenaline solutionwas injected at the planned incision site. Additional anaesthesia wasprovided, if required, by regional nerve block or subcutaneous injectionof lidocaine/adrenaline at the proposed implantation site. A dermalincision 1-2 mm in length was made with a scalpel (11 blade) or sharpinjection needle to facilitate transdermal insertion of a 16 Gblunt-tipped cannula for administration of the gel into thesubcutaneous, submuscular or supraperiostal adipose tissue. The gel wasinjected in small aliquots throughout the area requiring augmentation,rather than as a single bolus, by manipulating the cannula into adifferent tract after each injection, using a tunnelling technique. Amaximum of 10 ml (5 syringes) of gel was administered at each treatmentsession to a maximum of 3 separate anatomical sites (chin and bothcheeks). On completion of the injection, the treatment area was massagedto conform to the contour of the surrounding tissue and, if necessary,ice was applied briefly to reduce any swelling.

Satisfactory cheek and/or chin augmentation for at least 3 months wasobtained by the method. In particular, deep subcutaneous injection andsupraperiostal injection further prevented migration of the implant.

Example 3

Preparation of Gel Particles of Non-Animal Stabilized Hyaluronic Acidwith Longer Duration

Ten grams of hyaluronic acid prepared by fermentation of Streptococcuswas dissolved in 100 ml of 1% NaOH, pH>9. Cross-linking agent in theform of 1,4-butanediol diglycidyl ether was added to a concentration of0.2%. The solution was incubated at 40° C. for 4 h.

The alkaline gel was divided in two portions, which were individuallydiluted with an acidic water solution to reach neutral pH under mixing,yielding final hyaluronic acid concentrations of 20 mg/ml and 25 mg/ml,respectively. The gels were incubated for 12 h at 70° C. and cooled toroom temperature. The two gel portions were combined and mashed to thefinal particle size, approximately 3-4 mm.

Example 4

Breast Tissue Augmentation

Women with small breasts were injected with a gel obtainable e.g. by themethod of example 3. Each breast received 100 ml gel implanted under theglandular region, just on the pectoralis muscle, using a blunt 12 Gneedle in small aliquots. Care was taken not to disturb the naturaltissue. Twelve months following implantation, the breasts were still ingood shape with thin nodular implants. The implant did not blur theanalysis of mammography.

One woman had changed her mind about having gels in her breasts andrequested for removal of the implant. A blunt cannulae (12 G) was usedto suck back the gel. Almost all of the implant was aspirated as a cleartransparent gel fluid. Analysis showed that the gel maintained itsvolume but was slightly lower in concentration as compared to theinitial concentration (75% of initial), indicating an implant durationof about 2-3 years.

1. A soft tissue augmentation implant comprising particles of across-linked hyaluronic acid, wherein said cross-linked hyaluronic acidis cross-linked with a cross-linking agent, wherein a major volume ofsaid particles are injectable gel particles having a size, whensubjected to a physiological salt solution, in the range of from 1.5 to5 MM.
 2. A soft tissue augmentation implant according to claim 1,wherein said size is in the range of from 1.5 to 2.5 mm.
 3. A softtissue augmentation implant according to claim 1, wherein said size isin the range of from 2.5 to 5 mm.
 4. A method of soft tissueaugmentation in a mammal, including man, comprising subepidermaladministration at a site in said mammal where soft tissue augmentationis desirable, of an implant comprising injectable gel particles of aviscoelastic medium, a major volume of said particles having a size,when subjected to a physiological salt solution, in the range of from 1to 5 mm.
 5. A method according to claim 4, wherein said administrationis subcutaneous administration.
 6. A method according to claim 4,wherein said administration is submuscular administration.
 7. A methodaccording to claim 4, wherein said administration is supraperiostaladministration.
 8. A method according to claim 4, wherein said size isin the range of from 1 to 2.5 mm.
 9. A method according to claims 4,wherein said size is in the range of from 2.5 to 5 mm.
 10. A methodaccording to claim 8, wherein said site of soft tissue augmentation isselected from facial tissue and other tissues covered by exposed skin.11. A method according to claim 4, wherein said administration is asingle administration.
 12. A method according to claim 4, wherein saidadministration is a multiple-layer administration.
 13. A soft tissueaugmentation implant according to claim 1, wherein said cross-linkingagent is selected from the group consisting of aldehydes, epoxides,polyaziridyl compounds, glycidyl ethers, and divinylsulfones.
 14. A softtissue augmentation implant according to claim 13, wherein saidcross-linking agent is an epoxide.
 15. A soft tissue augmentationimplant according to claim 14, wherein said cross-linking agent is1,4-butanediol diglycidyl ether.
 16. A soft tissue augmentation implantaccording to claim 1, wherein the concentration of the cross-linkedhyaluronic acid in said gel particles, when subjected to a physiologicalsalt solution, is in the range of from 15 to 30 mg/ml.
 17. A soft tissueaugmentation implant according to claim 16, wherein the concentration ofthe cross-linked hyaluronic acid in said gel particles, when subjectedto a physiological salt solution, is in the range of from 22 to 30mg/ml.
 18. A soft tissue augmentation implant according to claim 16,wherein the concentration of the cross-linked hyaluronic acid in saidgel particles, when subjected to a physiological salt solution, is inthe range of from 20 to 25 mg/ml.
 19. A soft tissue augmentation implantaccording to claim 1, which are injectable through a 20 gauge or largerneedle by application of a pressure of 15-50 N.
 20. A soft tissueaugmentation implant according to claim 1, wherein removal of theimplant is facilitated compared to an implant with smaller particles.